The aging phenotype is due to the complex interaction of genetic, epigenetic, and environmental factors. Changes in the epigenetic control of gene expression may be one of the central mechanisms by which aging predisposes to many age-related diseases. In light of the above, epigenetic alterations may play a role in extension of lifespan and incidence of age-related degenerative diseases. Here we wish to systematically assess the contribution of epigenetic changes (i.e. a set of reversible, heritable changes in gene function or other cell phenotype that occurs without a change in DNA sequence), and in particular DNA methylation status to the aging process and phenotype. Absolute, altered methylation sites increases with increasing age, such that it can serve as marker for chronological age. We hypothesize that subjects with exceptional longevity may exhibit methylation at sites distinct from controls, in which case the absolute number of altered methylation sites will not change with age or even decrease as those subjects approach the end of their life. Thus, we predict that distinct DNA methylation sites may have a role in altering epigenetic profile in centenarians compared to people at younger ages. To determine if aging is associated with alterations in genome-wide methylation patterns and if individuals with exceptional longevity are changing, we will employ a novel technology to probe this new epigenetic hallmark for exceptional longevity in a population of subjects between ages 60- 110. We propose a combination of large-scale genomic studies (genome-wide cytosine methylation assay-HELPtag) to identify the most distinctive epigenetic loci (i.e. those with the greatest differences in methylation status). We will then perform Multi-locus validation using a rapid, sensitive, accurate and quantitative technique to analyze methylation patterns using the high throughput Sequenom MassARRAY platform. We will examine the association between epigenetic loci and favorable biological phenotypes in addition to age- related diseases, including cardiovascular diseases, metabolic syndrome and cognitive function in offspring of centenarian and age-match control. Finally we will establish expression patterns (qRT-PCR) of the candidate epigenetic loci identified in Aims 1 and 2 as well as specific genes (such as DMNTs and tumor suppressor genes) in blood cells of subjects among the initially screened population. Gene expression levels for all candidate and activity (for gene with pivotal role) will then be compared between the analyzed groups to demonstrate the impact of specific epigenetic loci on those genes and of genes in regulatory regions to establish possible mechanistic links between altered methylation with age and healthy life span. These studies will further our understanding of the complex process of aging and will identify loci that when altered epigenetically, have important ramifications for age-related diseases and therefore, life span. The data from the above research will lead to future investigations to determine if epigenetic differences can be observed longitudinally between offspring of centenarians and offspring of parents with usual life span, and whether differences predict the occurrence of age-related diseases. PUBLIC HEALTH RELEVANCE: Genetic and environmental factors play a crucial role in determining disease risk and life span. Recently, epigenetics has emerged as an important factor in the control of gene expression and disease risk. We hypothesize that methylation changes associated with aging are one of the central mechanisms by which aging predisposes to many age-related diseases, and may be also a marker for healthy life span. This research will further our understanding of the biological process to aging by employing un-bias approach to identifying loci that when altered epigenetically have important ramifications for age-related diseases and lifespan. Validating the genes whose function is modulated epigenetically could lead to interventions to delay or even prevent development of age-associated diseases.